The importance in eukaryotes of polyphospho-myo-inositides and polyphospho-myo-inositols in mediating cellular responses to specific extracellular signals is well established. In addition, myo-inositol is found in other important, more complex molecules such as glycosyl phosphatidylinositol (GPI) -anchored proteins, where it may also participate in signaling cascades. It is becoming increasingly apparent that non-myoinositol-containing, isomeric phosphoinositides are more common than once believed, with (1D )chiro-and/or scyllo-inositol-containing phosphatidylinositols having been reported in mammals, plants, and a lower eukaryote. Given the importance of understanding the potential of non-myo-inositol containing lipids and second messengers in transmembrane and intracellular signaling, it is imperative to examine the biochemistry of these molecules. Using the technique of radioisotopic labeling in conjunction with kinetic analyses, chromatography, electrophoresis, and other methods, the applicant will (1) characterize the transport kinetics of three inositol isomers, (2) examine the utilization of these isomers in the biosynthesis of phosphoinositides, phosphoinositols, and GPI-anchored proteins, (3) characterize the utilization of inositol isomers by phosphatidylinositol synthase, and (4) determine the susceptibility of non-myo-inositolcontaining phosphatidylinositol to degradation by seven common phospholipases. By understanding the basic mechanisms of inositol isomer uptake, utilization, biosynthesis, and degradation, the foundation is laid for accomplishing the long term goals of this project, which include an examination of the structure/function relationships among the isomeric inositol lipids, the influence these lipids exert on the biophysical properties of membranes, and the role non-myo-inositol second messengers play in signaling cascades.